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cux1 p110  (Addgene inc)


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    Addgene inc cux1 p110
    Expression profiling was performed on microdissected tumor cells from 17 mammary tumors that developed in <t>MMTV-p110</t> or p75 <t>CUX1</t> transgenic mice, including 8 solid carcinomas (p110-416, p110-314, p110-360, p75-534, p75-150, p110-143, p75-129, p110-225), 4 adenosquamous carcinomas (p75-101, p75-94, p75-80, p75-47), 1 adenoma (p110-98), 2 adenomyoepitheliomas (p75-90, p110-374), and 1 adenocarcinoma (p75-216). The complete expression profiles are available on the Gene Expression Omnibus (GEO) repository under accession number GSE54804. The figure shows a heatmap of mammary tumors grouped according to an unsupervised hierarchical clustering of Wnt gene expression. Expression of Glis1 , as well as Vim , Cdh1 , Cdh2 , Snai1 , Snai2 , Ocln and Twist1 in each tumor is shown below. The cluster of “high-Wnt” tumors is boxed in yellow and the cluster of “low-Wnt” tumors is boxed in blue. Each column represents one tumor, with the transgene (p110 or p75 CUX1) and mouse ID number indicated below.
    Cux1 P110, supplied by Addgene inc, used in various techniques. Bioz Stars score: 85/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/product/cux1+p110/pLXSN+p110+CUX1+(Plasmid+%2390471)/pmc04197442-65-35-29
    Average 85 stars, based on 1 article reviews
    cux1 p110 - by Bioz Stars, 2026-09
    85/100 stars

    Images

    1) Product Images from "Autocrine Activation of the Wnt/β-Catenin Pathway by CUX1 and GLIS1 in Breast Cancers"

    Article Title: Autocrine Activation of the Wnt/β-Catenin Pathway by CUX1 and GLIS1 in Breast Cancers

    Journal: Biology Open

    doi: 10.1242/bio.20148193

    Expression profiling was performed on microdissected tumor cells from 17 mammary tumors that developed in MMTV-p110 or p75 CUX1 transgenic mice, including 8 solid carcinomas (p110-416, p110-314, p110-360, p75-534, p75-150, p110-143, p75-129, p110-225), 4 adenosquamous carcinomas (p75-101, p75-94, p75-80, p75-47), 1 adenoma (p110-98), 2 adenomyoepitheliomas (p75-90, p110-374), and 1 adenocarcinoma (p75-216). The complete expression profiles are available on the Gene Expression Omnibus (GEO) repository under accession number GSE54804. The figure shows a heatmap of mammary tumors grouped according to an unsupervised hierarchical clustering of Wnt gene expression. Expression of Glis1 , as well as Vim , Cdh1 , Cdh2 , Snai1 , Snai2 , Ocln and Twist1 in each tumor is shown below. The cluster of “high-Wnt” tumors is boxed in yellow and the cluster of “low-Wnt” tumors is boxed in blue. Each column represents one tumor, with the transgene (p110 or p75 CUX1) and mouse ID number indicated below.
    Figure Legend Snippet: Expression profiling was performed on microdissected tumor cells from 17 mammary tumors that developed in MMTV-p110 or p75 CUX1 transgenic mice, including 8 solid carcinomas (p110-416, p110-314, p110-360, p75-534, p75-150, p110-143, p75-129, p110-225), 4 adenosquamous carcinomas (p75-101, p75-94, p75-80, p75-47), 1 adenoma (p110-98), 2 adenomyoepitheliomas (p75-90, p110-374), and 1 adenocarcinoma (p75-216). The complete expression profiles are available on the Gene Expression Omnibus (GEO) repository under accession number GSE54804. The figure shows a heatmap of mammary tumors grouped according to an unsupervised hierarchical clustering of Wnt gene expression. Expression of Glis1 , as well as Vim , Cdh1 , Cdh2 , Snai1 , Snai2 , Ocln and Twist1 in each tumor is shown below. The cluster of “high-Wnt” tumors is boxed in yellow and the cluster of “low-Wnt” tumors is boxed in blue. Each column represents one tumor, with the transgene (p110 or p75 CUX1) and mouse ID number indicated below.

    Techniques Used: Expressing, Transgenic Assay, Gene Expression

    Cell lines were established from two mammary tumors that developed in transgenic mice carrying a MMTV-p75 CUX1 transgene: an adenosquamous carcinoma from mouse #p75-80 and a solid carcinoma from mouse #p75-534. The two cell lines were analyzed for Wnt gene expression (A), nuclear β-catenin expression (B), and TCF/β-catenin transcriptional activity (C). (A) mRNA levels of Wnt ligand genes were measured by RT-qPCR. Results of triplicate experiments are shown. * P Value<0.05 on a Welch corrected T test. Und: Undetected. (B) Cells were analyzed by indirect immunofluorescence using antibodies specific for the active, non-phosphorylated form of β-Catenin. Histogram shows the relative mean nuclear signal for β-Catenin as measured using the ImageJ software. The value for p75-534 was set to 1. * P Value<0.05 on a Welch corrected T test. A representative image of each cell line is shown on the right. Scale bar: 20µm. (C) The TOP/FOP luciferase reporter assay was performed in the p75-534 and p75-80 mammary tumor cell lines. In addition, p75-80 cells were cotransfected with a vector expressing either sFRP1, sFRP2 or SOST or an empty vector. Where indicated, niclosamide was added to the cells at the time of transfection. Results of 3 independent transfections are shown. * P Value<0.05 on a Welch corrected T test. A schematic representation of the reporter constructs is shown.
    Figure Legend Snippet: Cell lines were established from two mammary tumors that developed in transgenic mice carrying a MMTV-p75 CUX1 transgene: an adenosquamous carcinoma from mouse #p75-80 and a solid carcinoma from mouse #p75-534. The two cell lines were analyzed for Wnt gene expression (A), nuclear β-catenin expression (B), and TCF/β-catenin transcriptional activity (C). (A) mRNA levels of Wnt ligand genes were measured by RT-qPCR. Results of triplicate experiments are shown. * P Value<0.05 on a Welch corrected T test. Und: Undetected. (B) Cells were analyzed by indirect immunofluorescence using antibodies specific for the active, non-phosphorylated form of β-Catenin. Histogram shows the relative mean nuclear signal for β-Catenin as measured using the ImageJ software. The value for p75-534 was set to 1. * P Value<0.05 on a Welch corrected T test. A representative image of each cell line is shown on the right. Scale bar: 20µm. (C) The TOP/FOP luciferase reporter assay was performed in the p75-534 and p75-80 mammary tumor cell lines. In addition, p75-80 cells were cotransfected with a vector expressing either sFRP1, sFRP2 or SOST or an empty vector. Where indicated, niclosamide was added to the cells at the time of transfection. Results of 3 independent transfections are shown. * P Value<0.05 on a Welch corrected T test. A schematic representation of the reporter constructs is shown.

    Techniques Used: Transgenic Assay, Gene Expression, Expressing, Activity Assay, Quantitative RT-PCR, Immunofluorescence, Software, Luciferase, Reporter Assay, Plasmid Preparation, Transfection, Construct

    CUX1 specific or scrambled siRNA were transfected in a panel of 6 human cancer cell lines from breast (A), ovarian (B) and lung (C) cancers. 3 days later total mRNA was isolated and quantitative RT-PCR was performed using HPRT1 mRNA as a control. The values represent fold difference in mRNA expression between cells treated with CUX1 or scrambled siRNA. (Und: undetected). * P Value<0.05 on a Welch corrected T test.
    Figure Legend Snippet: CUX1 specific or scrambled siRNA were transfected in a panel of 6 human cancer cell lines from breast (A), ovarian (B) and lung (C) cancers. 3 days later total mRNA was isolated and quantitative RT-PCR was performed using HPRT1 mRNA as a control. The values represent fold difference in mRNA expression between cells treated with CUX1 or scrambled siRNA. (Und: undetected). * P Value<0.05 on a Welch corrected T test.

    Techniques Used: Transfection, Isolation, Quantitative RT-PCR, Control, Expressing

    (A) HEK293 cells were infected with retroviruses to establish cells stably carrying a vector that expresses p110 CUX1 or nothing (vector). Nuclear and cytoplasmic extracts were analyzed by immunoblotting with the indicated antibodies. (B) The TOP or FOP luciferase reporters were introduced into HEK293 cells together with 500 ng of a vector expressing p110 CUX1 or an empty vector. Luciferase activity was measured 36 hours after transfection. Where indicated, a vector expressing either sFRP1, sFRP2, SOST or DKK1 was transfected as well, or niclosamide or IWP-2 was added to the cells at the time of transfection. Results of 3 independent transfections are shown. * P Value<0.05 on a Welch corrected T test. (C) Hs578t and MCF7 breast cancer cells were stably infected with either a lentivirus expressing p110 CUX1 or an empty lentivirus (vector). Nuclear and cytoplasmic extracts were analyzed by immunoblotting with the indicated antibodies. (D) The TOP or FOP luciferase reporters were introduced into Hs578t and MCF7 cells together with 500 ng of a vector expressing p110 CUX1 or an empty vector. Luciferase activity was measured 36 hours after transfection.* P Value<0.05 on a Welch corrected T test.
    Figure Legend Snippet: (A) HEK293 cells were infected with retroviruses to establish cells stably carrying a vector that expresses p110 CUX1 or nothing (vector). Nuclear and cytoplasmic extracts were analyzed by immunoblotting with the indicated antibodies. (B) The TOP or FOP luciferase reporters were introduced into HEK293 cells together with 500 ng of a vector expressing p110 CUX1 or an empty vector. Luciferase activity was measured 36 hours after transfection. Where indicated, a vector expressing either sFRP1, sFRP2, SOST or DKK1 was transfected as well, or niclosamide or IWP-2 was added to the cells at the time of transfection. Results of 3 independent transfections are shown. * P Value<0.05 on a Welch corrected T test. (C) Hs578t and MCF7 breast cancer cells were stably infected with either a lentivirus expressing p110 CUX1 or an empty lentivirus (vector). Nuclear and cytoplasmic extracts were analyzed by immunoblotting with the indicated antibodies. (D) The TOP or FOP luciferase reporters were introduced into Hs578t and MCF7 cells together with 500 ng of a vector expressing p110 CUX1 or an empty vector. Luciferase activity was measured 36 hours after transfection.* P Value<0.05 on a Welch corrected T test.

    Techniques Used: Infection, Stable Transfection, Plasmid Preparation, Western Blot, Luciferase, Expressing, Activity Assay, Transfection

    (A) Heatmap of a human breast tumor dataset sorted according to WNT genes expression using the BreSAT algorithm. Expression of CUX1 , GLIS1 , VIM , TWIST1 , SNAI1 , CDH2 , SNAI2 , OCLN and CDH1 in each tumor is shown below. (B) CUX1 and GLIS1 expression in the top 25% and bottom 25% samples sorted according to WNT genes expression. * indicates p<0.05, ** <0.01, *** <0.001 on a Welch-corrected student's T test. (C) CDH1 , OCLN , CDH2 , SNAI1 , SNAI2 , TWIST1 and VIM expression in the top 25% and bottom 25% samples sorted according to Wnt genes expression. * indicates p<0.05, ** <0.01, *** <0.001 on a Welch-corrected student's T test.
    Figure Legend Snippet: (A) Heatmap of a human breast tumor dataset sorted according to WNT genes expression using the BreSAT algorithm. Expression of CUX1 , GLIS1 , VIM , TWIST1 , SNAI1 , CDH2 , SNAI2 , OCLN and CDH1 in each tumor is shown below. (B) CUX1 and GLIS1 expression in the top 25% and bottom 25% samples sorted according to WNT genes expression. * indicates p<0.05, ** <0.01, *** <0.001 on a Welch-corrected student's T test. (C) CDH1 , OCLN , CDH2 , SNAI1 , SNAI2 , TWIST1 and VIM expression in the top 25% and bottom 25% samples sorted according to Wnt genes expression. * indicates p<0.05, ** <0.01, *** <0.001 on a Welch-corrected student's T test.

    Techniques Used: Expressing

    We established populations of MCF10A cells stably carrying lentiviral vectors that express nothing (vector), GLIS1, p110 CUX1, or both GLIS1 and p110 CUX1. (A) Wnt gene mRNA levels were measured by RT-qPCR. Und: Undetected. * P Value<0.05 on a Welch corrected T test when comparing the p110+Glis1 samples vs. each of the other 3. (B) β-Catenin protein levels were measured by immunofluorescence using antibodies specific for the active, non-phosphorylated form of the protein. Histogram shows the mean nuclear signal for β-Catenin for at least 50 cells per condition. * P Value<0.05 on a Welch corrected T test. A representative image of each condition is shown below. Scale bar: 20µm. (C) β-Catenin protein levels were measured by immunoblotting using antibodies specific for the active, non-phosphorylated form of the protein. γ-Tubulin is shown as a loading control. (D) MCF10A cells were transiently transfected with either the TOP of the FOP luciferase reporter, a vector expressing p110 CUX1 or an empty vector, and a vector expressing GLIS1 or the corresponding empty vector. Luciferase activity was measured in 3 independent transfections. * P Value<0.05 on a Welch corrected T test. (E) MCF10A cells ectopically expressing GLIS1, p110 CUX1 or both were imaged by time-lapse video microscopy and their speed of migration was quantified. The average of 3 independent experiments is shown with error bars representing standard error. ** P Value<0.01, *** <0.001 on 3 combined Welch corrected T tests using the Fisher method. (F) Time-lapse video microscopy was carried out as in E and distance migrated from the point of origin was quantified. (G) MCF10A cells overexpressing GLIS1 and p110 CUX1 alone or both were subjected to inverted Boyden chamber invasion assays. The percentage of cells able to migrate more than 20 µm into the matrix is shown for each line. The average of 3 independent experiments is shown. (H) Confocal Z-Stacks acquired 10 µm apart are aligned to demonstrate invasion of MCF10A-derivative cells into the matrix. Representative Z-stack for each condition is displayed. The leftmost image represents cells at the surface of the matrix (Depth = 0).
    Figure Legend Snippet: We established populations of MCF10A cells stably carrying lentiviral vectors that express nothing (vector), GLIS1, p110 CUX1, or both GLIS1 and p110 CUX1. (A) Wnt gene mRNA levels were measured by RT-qPCR. Und: Undetected. * P Value<0.05 on a Welch corrected T test when comparing the p110+Glis1 samples vs. each of the other 3. (B) β-Catenin protein levels were measured by immunofluorescence using antibodies specific for the active, non-phosphorylated form of the protein. Histogram shows the mean nuclear signal for β-Catenin for at least 50 cells per condition. * P Value<0.05 on a Welch corrected T test. A representative image of each condition is shown below. Scale bar: 20µm. (C) β-Catenin protein levels were measured by immunoblotting using antibodies specific for the active, non-phosphorylated form of the protein. γ-Tubulin is shown as a loading control. (D) MCF10A cells were transiently transfected with either the TOP of the FOP luciferase reporter, a vector expressing p110 CUX1 or an empty vector, and a vector expressing GLIS1 or the corresponding empty vector. Luciferase activity was measured in 3 independent transfections. * P Value<0.05 on a Welch corrected T test. (E) MCF10A cells ectopically expressing GLIS1, p110 CUX1 or both were imaged by time-lapse video microscopy and their speed of migration was quantified. The average of 3 independent experiments is shown with error bars representing standard error. ** P Value<0.01, *** <0.001 on 3 combined Welch corrected T tests using the Fisher method. (F) Time-lapse video microscopy was carried out as in E and distance migrated from the point of origin was quantified. (G) MCF10A cells overexpressing GLIS1 and p110 CUX1 alone or both were subjected to inverted Boyden chamber invasion assays. The percentage of cells able to migrate more than 20 µm into the matrix is shown for each line. The average of 3 independent experiments is shown. (H) Confocal Z-Stacks acquired 10 µm apart are aligned to demonstrate invasion of MCF10A-derivative cells into the matrix. Representative Z-stack for each condition is displayed. The leftmost image represents cells at the surface of the matrix (Depth = 0).

    Techniques Used: Stable Transfection, Plasmid Preparation, Quantitative RT-PCR, Immunofluorescence, Western Blot, Control, Transfection, Luciferase, Expressing, Activity Assay, Microscopy, Migration

    We established populations of Hs578t cells stably carrying lentiviral vectors that express nothing (vector), GLIS1, p110 CUX1, or both GLIS1 and p110 CUX1. (A) Wnt gene mRNA levels were measured by RT-qPCR. Und: Undetected. * P Value<0.05 on a Welch corrected T test. (B) β-Catenin protein levels were measured by indirect immunofluorescence using antibodies specific for the active, non-phosphorylated form of the protein. Histogram shows the mean nuclear signal for β-Catenin for at least 50 cells per condition. * P Value<0.05 on a Welch corrected T test. A representative image of each condition is shown below. Scale bar: 20µm. (C) Hs578t cells were transiently transfected with either the TOP of the FOP luciferase reporter, and suboptimal amounts (50 ng) of effector plasmids coding for nothing (−), p110 CUX1 or GLIS1. Luciferase activity was measured in 3 independent transfections. * P Value<0.05 on a Welch corrected T test. (D) Hs578t cells ectopically expressing GLIS1, p110 CUX1 or both were imaged by time-lapse video microscopy and their speed of migration was quantified. The average of 3 independent experiments is shown with error bars representing standard error. ** P Value<0.01, *** <0.001 on a Welch corrected T test. (E) Time-lapse video microscopy was carried out as in D and distance migrated from the point of origin was quantified.
    Figure Legend Snippet: We established populations of Hs578t cells stably carrying lentiviral vectors that express nothing (vector), GLIS1, p110 CUX1, or both GLIS1 and p110 CUX1. (A) Wnt gene mRNA levels were measured by RT-qPCR. Und: Undetected. * P Value<0.05 on a Welch corrected T test. (B) β-Catenin protein levels were measured by indirect immunofluorescence using antibodies specific for the active, non-phosphorylated form of the protein. Histogram shows the mean nuclear signal for β-Catenin for at least 50 cells per condition. * P Value<0.05 on a Welch corrected T test. A representative image of each condition is shown below. Scale bar: 20µm. (C) Hs578t cells were transiently transfected with either the TOP of the FOP luciferase reporter, and suboptimal amounts (50 ng) of effector plasmids coding for nothing (−), p110 CUX1 or GLIS1. Luciferase activity was measured in 3 independent transfections. * P Value<0.05 on a Welch corrected T test. (D) Hs578t cells ectopically expressing GLIS1, p110 CUX1 or both were imaged by time-lapse video microscopy and their speed of migration was quantified. The average of 3 independent experiments is shown with error bars representing standard error. ** P Value<0.01, *** <0.001 on a Welch corrected T test. (E) Time-lapse video microscopy was carried out as in D and distance migrated from the point of origin was quantified.

    Techniques Used: Stable Transfection, Plasmid Preparation, Quantitative RT-PCR, Immunofluorescence, Transfection, Luciferase, Activity Assay, Expressing, Microscopy, Migration

    Related Articles

    Reporter Assay:

    Article Title: Autocrine Activation of the Wnt/β-Catenin Pathway by CUX1 and GLIS1 in Breast Cancers
    Article Snippet: .. For TOP/FOP reporter assay, Cells plated at 8×10 4 per well in 12-well plates were cotransfected with 0.25 μg of either the TOP 8× or FOP 8× reporter plasmids (Addgene) and 0.5 μg of either CUX1 p110 or empty vector as effector. .. As a control for transfection efficiency, the β-galactosidase protein (Sigma) was included in the transfection mix and the luciferase activity was normalized based on β-galactosidase activity.



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    Image Search Results


    Fig. 4. NE co-localizes and interacts with p200 CUX1. (A) Expression levels of

    Journal: Life sciences

    Article Title: Neutrophil elastase-mediated proteolysis of the tumor suppressor p200 CUX1 promotes cell proliferation and inhibits cell differentiation in APL.

    doi: 10.1016/j.lfs.2019.117229

    Figure Lengend Snippet: Fig. 4. NE co-localizes and interacts with p200 CUX1. (A) Expression levels of

    Article Snippet: Primary antibodies were purchased from the following sources: against human NE (no. ab131260) from Abcam (UK); against p200 CUX1 (no. 81557), P21 Waf1/Cip1 (no. 2947), P27Kip1 (no. 3686), and C/EBPα (no. 8178) from Cell Signaling Technology (CST, USA); against p110/p75 CUX1 (no. TA890108) from Origene Technologies (Origene, USA); against C/EBPβ (no. sc-137146) from Santa Cruz Biotechnology (Santa Cruz, USA).

    Techniques: Expressing

    Fig. 5. NE effectively hydrolyzes p200 CUX1. (A) NE overexpression decreased

    Journal: Life sciences

    Article Title: Neutrophil elastase-mediated proteolysis of the tumor suppressor p200 CUX1 promotes cell proliferation and inhibits cell differentiation in APL.

    doi: 10.1016/j.lfs.2019.117229

    Figure Lengend Snippet: Fig. 5. NE effectively hydrolyzes p200 CUX1. (A) NE overexpression decreased

    Article Snippet: Primary antibodies were purchased from the following sources: against human NE (no. ab131260) from Abcam (UK); against p200 CUX1 (no. 81557), P21 Waf1/Cip1 (no. 2947), P27Kip1 (no. 3686), and C/EBPα (no. 8178) from Cell Signaling Technology (CST, USA); against p110/p75 CUX1 (no. TA890108) from Origene Technologies (Origene, USA); against C/EBPβ (no. sc-137146) from Santa Cruz Biotechnology (Santa Cruz, USA).

    Techniques: Over Expression

    Fig. 6. p200 CUX1 overexpression reverses the effects of NE on cell proliferation

    Journal: Life sciences

    Article Title: Neutrophil elastase-mediated proteolysis of the tumor suppressor p200 CUX1 promotes cell proliferation and inhibits cell differentiation in APL.

    doi: 10.1016/j.lfs.2019.117229

    Figure Lengend Snippet: Fig. 6. p200 CUX1 overexpression reverses the effects of NE on cell proliferation

    Article Snippet: Primary antibodies were purchased from the following sources: against human NE (no. ab131260) from Abcam (UK); against p200 CUX1 (no. 81557), P21 Waf1/Cip1 (no. 2947), P27Kip1 (no. 3686), and C/EBPα (no. 8178) from Cell Signaling Technology (CST, USA); against p110/p75 CUX1 (no. TA890108) from Origene Technologies (Origene, USA); against C/EBPβ (no. sc-137146) from Santa Cruz Biotechnology (Santa Cruz, USA).

    Techniques: Over Expression

    Expression profiling was performed on microdissected tumor cells from 17 mammary tumors that developed in MMTV-p110 or p75 CUX1 transgenic mice, including 8 solid carcinomas (p110-416, p110-314, p110-360, p75-534, p75-150, p110-143, p75-129, p110-225), 4 adenosquamous carcinomas (p75-101, p75-94, p75-80, p75-47), 1 adenoma (p110-98), 2 adenomyoepitheliomas (p75-90, p110-374), and 1 adenocarcinoma (p75-216). The complete expression profiles are available on the Gene Expression Omnibus (GEO) repository under accession number GSE54804. The figure shows a heatmap of mammary tumors grouped according to an unsupervised hierarchical clustering of Wnt gene expression. Expression of Glis1 , as well as Vim , Cdh1 , Cdh2 , Snai1 , Snai2 , Ocln and Twist1 in each tumor is shown below. The cluster of “high-Wnt” tumors is boxed in yellow and the cluster of “low-Wnt” tumors is boxed in blue. Each column represents one tumor, with the transgene (p110 or p75 CUX1) and mouse ID number indicated below.

    Journal: Biology Open

    Article Title: Autocrine Activation of the Wnt/β-Catenin Pathway by CUX1 and GLIS1 in Breast Cancers

    doi: 10.1242/bio.20148193

    Figure Lengend Snippet: Expression profiling was performed on microdissected tumor cells from 17 mammary tumors that developed in MMTV-p110 or p75 CUX1 transgenic mice, including 8 solid carcinomas (p110-416, p110-314, p110-360, p75-534, p75-150, p110-143, p75-129, p110-225), 4 adenosquamous carcinomas (p75-101, p75-94, p75-80, p75-47), 1 adenoma (p110-98), 2 adenomyoepitheliomas (p75-90, p110-374), and 1 adenocarcinoma (p75-216). The complete expression profiles are available on the Gene Expression Omnibus (GEO) repository under accession number GSE54804. The figure shows a heatmap of mammary tumors grouped according to an unsupervised hierarchical clustering of Wnt gene expression. Expression of Glis1 , as well as Vim , Cdh1 , Cdh2 , Snai1 , Snai2 , Ocln and Twist1 in each tumor is shown below. The cluster of “high-Wnt” tumors is boxed in yellow and the cluster of “low-Wnt” tumors is boxed in blue. Each column represents one tumor, with the transgene (p110 or p75 CUX1) and mouse ID number indicated below.

    Article Snippet: For TOP/FOP reporter assay, Cells plated at 8×10 4 per well in 12-well plates were cotransfected with 0.25 μg of either the TOP 8× or FOP 8× reporter plasmids (Addgene) and 0.5 μg of either CUX1 p110 or empty vector as effector.

    Techniques: Expressing, Transgenic Assay, Gene Expression

    Cell lines were established from two mammary tumors that developed in transgenic mice carrying a MMTV-p75 CUX1 transgene: an adenosquamous carcinoma from mouse #p75-80 and a solid carcinoma from mouse #p75-534. The two cell lines were analyzed for Wnt gene expression (A), nuclear β-catenin expression (B), and TCF/β-catenin transcriptional activity (C). (A) mRNA levels of Wnt ligand genes were measured by RT-qPCR. Results of triplicate experiments are shown. * P Value<0.05 on a Welch corrected T test. Und: Undetected. (B) Cells were analyzed by indirect immunofluorescence using antibodies specific for the active, non-phosphorylated form of β-Catenin. Histogram shows the relative mean nuclear signal for β-Catenin as measured using the ImageJ software. The value for p75-534 was set to 1. * P Value<0.05 on a Welch corrected T test. A representative image of each cell line is shown on the right. Scale bar: 20µm. (C) The TOP/FOP luciferase reporter assay was performed in the p75-534 and p75-80 mammary tumor cell lines. In addition, p75-80 cells were cotransfected with a vector expressing either sFRP1, sFRP2 or SOST or an empty vector. Where indicated, niclosamide was added to the cells at the time of transfection. Results of 3 independent transfections are shown. * P Value<0.05 on a Welch corrected T test. A schematic representation of the reporter constructs is shown.

    Journal: Biology Open

    Article Title: Autocrine Activation of the Wnt/β-Catenin Pathway by CUX1 and GLIS1 in Breast Cancers

    doi: 10.1242/bio.20148193

    Figure Lengend Snippet: Cell lines were established from two mammary tumors that developed in transgenic mice carrying a MMTV-p75 CUX1 transgene: an adenosquamous carcinoma from mouse #p75-80 and a solid carcinoma from mouse #p75-534. The two cell lines were analyzed for Wnt gene expression (A), nuclear β-catenin expression (B), and TCF/β-catenin transcriptional activity (C). (A) mRNA levels of Wnt ligand genes were measured by RT-qPCR. Results of triplicate experiments are shown. * P Value<0.05 on a Welch corrected T test. Und: Undetected. (B) Cells were analyzed by indirect immunofluorescence using antibodies specific for the active, non-phosphorylated form of β-Catenin. Histogram shows the relative mean nuclear signal for β-Catenin as measured using the ImageJ software. The value for p75-534 was set to 1. * P Value<0.05 on a Welch corrected T test. A representative image of each cell line is shown on the right. Scale bar: 20µm. (C) The TOP/FOP luciferase reporter assay was performed in the p75-534 and p75-80 mammary tumor cell lines. In addition, p75-80 cells were cotransfected with a vector expressing either sFRP1, sFRP2 or SOST or an empty vector. Where indicated, niclosamide was added to the cells at the time of transfection. Results of 3 independent transfections are shown. * P Value<0.05 on a Welch corrected T test. A schematic representation of the reporter constructs is shown.

    Article Snippet: For TOP/FOP reporter assay, Cells plated at 8×10 4 per well in 12-well plates were cotransfected with 0.25 μg of either the TOP 8× or FOP 8× reporter plasmids (Addgene) and 0.5 μg of either CUX1 p110 or empty vector as effector.

    Techniques: Transgenic Assay, Gene Expression, Expressing, Activity Assay, Quantitative RT-PCR, Immunofluorescence, Software, Luciferase, Reporter Assay, Plasmid Preparation, Transfection, Construct

    CUX1 specific or scrambled siRNA were transfected in a panel of 6 human cancer cell lines from breast (A), ovarian (B) and lung (C) cancers. 3 days later total mRNA was isolated and quantitative RT-PCR was performed using HPRT1 mRNA as a control. The values represent fold difference in mRNA expression between cells treated with CUX1 or scrambled siRNA. (Und: undetected). * P Value<0.05 on a Welch corrected T test.

    Journal: Biology Open

    Article Title: Autocrine Activation of the Wnt/β-Catenin Pathway by CUX1 and GLIS1 in Breast Cancers

    doi: 10.1242/bio.20148193

    Figure Lengend Snippet: CUX1 specific or scrambled siRNA were transfected in a panel of 6 human cancer cell lines from breast (A), ovarian (B) and lung (C) cancers. 3 days later total mRNA was isolated and quantitative RT-PCR was performed using HPRT1 mRNA as a control. The values represent fold difference in mRNA expression between cells treated with CUX1 or scrambled siRNA. (Und: undetected). * P Value<0.05 on a Welch corrected T test.

    Article Snippet: For TOP/FOP reporter assay, Cells plated at 8×10 4 per well in 12-well plates were cotransfected with 0.25 μg of either the TOP 8× or FOP 8× reporter plasmids (Addgene) and 0.5 μg of either CUX1 p110 or empty vector as effector.

    Techniques: Transfection, Isolation, Quantitative RT-PCR, Control, Expressing

    (A) HEK293 cells were infected with retroviruses to establish cells stably carrying a vector that expresses p110 CUX1 or nothing (vector). Nuclear and cytoplasmic extracts were analyzed by immunoblotting with the indicated antibodies. (B) The TOP or FOP luciferase reporters were introduced into HEK293 cells together with 500 ng of a vector expressing p110 CUX1 or an empty vector. Luciferase activity was measured 36 hours after transfection. Where indicated, a vector expressing either sFRP1, sFRP2, SOST or DKK1 was transfected as well, or niclosamide or IWP-2 was added to the cells at the time of transfection. Results of 3 independent transfections are shown. * P Value<0.05 on a Welch corrected T test. (C) Hs578t and MCF7 breast cancer cells were stably infected with either a lentivirus expressing p110 CUX1 or an empty lentivirus (vector). Nuclear and cytoplasmic extracts were analyzed by immunoblotting with the indicated antibodies. (D) The TOP or FOP luciferase reporters were introduced into Hs578t and MCF7 cells together with 500 ng of a vector expressing p110 CUX1 or an empty vector. Luciferase activity was measured 36 hours after transfection.* P Value<0.05 on a Welch corrected T test.

    Journal: Biology Open

    Article Title: Autocrine Activation of the Wnt/β-Catenin Pathway by CUX1 and GLIS1 in Breast Cancers

    doi: 10.1242/bio.20148193

    Figure Lengend Snippet: (A) HEK293 cells were infected with retroviruses to establish cells stably carrying a vector that expresses p110 CUX1 or nothing (vector). Nuclear and cytoplasmic extracts were analyzed by immunoblotting with the indicated antibodies. (B) The TOP or FOP luciferase reporters were introduced into HEK293 cells together with 500 ng of a vector expressing p110 CUX1 or an empty vector. Luciferase activity was measured 36 hours after transfection. Where indicated, a vector expressing either sFRP1, sFRP2, SOST or DKK1 was transfected as well, or niclosamide or IWP-2 was added to the cells at the time of transfection. Results of 3 independent transfections are shown. * P Value<0.05 on a Welch corrected T test. (C) Hs578t and MCF7 breast cancer cells were stably infected with either a lentivirus expressing p110 CUX1 or an empty lentivirus (vector). Nuclear and cytoplasmic extracts were analyzed by immunoblotting with the indicated antibodies. (D) The TOP or FOP luciferase reporters were introduced into Hs578t and MCF7 cells together with 500 ng of a vector expressing p110 CUX1 or an empty vector. Luciferase activity was measured 36 hours after transfection.* P Value<0.05 on a Welch corrected T test.

    Article Snippet: For TOP/FOP reporter assay, Cells plated at 8×10 4 per well in 12-well plates were cotransfected with 0.25 μg of either the TOP 8× or FOP 8× reporter plasmids (Addgene) and 0.5 μg of either CUX1 p110 or empty vector as effector.

    Techniques: Infection, Stable Transfection, Plasmid Preparation, Western Blot, Luciferase, Expressing, Activity Assay, Transfection

    (A) Heatmap of a human breast tumor dataset sorted according to WNT genes expression using the BreSAT algorithm. Expression of CUX1 , GLIS1 , VIM , TWIST1 , SNAI1 , CDH2 , SNAI2 , OCLN and CDH1 in each tumor is shown below. (B) CUX1 and GLIS1 expression in the top 25% and bottom 25% samples sorted according to WNT genes expression. * indicates p<0.05, ** <0.01, *** <0.001 on a Welch-corrected student's T test. (C) CDH1 , OCLN , CDH2 , SNAI1 , SNAI2 , TWIST1 and VIM expression in the top 25% and bottom 25% samples sorted according to Wnt genes expression. * indicates p<0.05, ** <0.01, *** <0.001 on a Welch-corrected student's T test.

    Journal: Biology Open

    Article Title: Autocrine Activation of the Wnt/β-Catenin Pathway by CUX1 and GLIS1 in Breast Cancers

    doi: 10.1242/bio.20148193

    Figure Lengend Snippet: (A) Heatmap of a human breast tumor dataset sorted according to WNT genes expression using the BreSAT algorithm. Expression of CUX1 , GLIS1 , VIM , TWIST1 , SNAI1 , CDH2 , SNAI2 , OCLN and CDH1 in each tumor is shown below. (B) CUX1 and GLIS1 expression in the top 25% and bottom 25% samples sorted according to WNT genes expression. * indicates p<0.05, ** <0.01, *** <0.001 on a Welch-corrected student's T test. (C) CDH1 , OCLN , CDH2 , SNAI1 , SNAI2 , TWIST1 and VIM expression in the top 25% and bottom 25% samples sorted according to Wnt genes expression. * indicates p<0.05, ** <0.01, *** <0.001 on a Welch-corrected student's T test.

    Article Snippet: For TOP/FOP reporter assay, Cells plated at 8×10 4 per well in 12-well plates were cotransfected with 0.25 μg of either the TOP 8× or FOP 8× reporter plasmids (Addgene) and 0.5 μg of either CUX1 p110 or empty vector as effector.

    Techniques: Expressing

    We established populations of MCF10A cells stably carrying lentiviral vectors that express nothing (vector), GLIS1, p110 CUX1, or both GLIS1 and p110 CUX1. (A) Wnt gene mRNA levels were measured by RT-qPCR. Und: Undetected. * P Value<0.05 on a Welch corrected T test when comparing the p110+Glis1 samples vs. each of the other 3. (B) β-Catenin protein levels were measured by immunofluorescence using antibodies specific for the active, non-phosphorylated form of the protein. Histogram shows the mean nuclear signal for β-Catenin for at least 50 cells per condition. * P Value<0.05 on a Welch corrected T test. A representative image of each condition is shown below. Scale bar: 20µm. (C) β-Catenin protein levels were measured by immunoblotting using antibodies specific for the active, non-phosphorylated form of the protein. γ-Tubulin is shown as a loading control. (D) MCF10A cells were transiently transfected with either the TOP of the FOP luciferase reporter, a vector expressing p110 CUX1 or an empty vector, and a vector expressing GLIS1 or the corresponding empty vector. Luciferase activity was measured in 3 independent transfections. * P Value<0.05 on a Welch corrected T test. (E) MCF10A cells ectopically expressing GLIS1, p110 CUX1 or both were imaged by time-lapse video microscopy and their speed of migration was quantified. The average of 3 independent experiments is shown with error bars representing standard error. ** P Value<0.01, *** <0.001 on 3 combined Welch corrected T tests using the Fisher method. (F) Time-lapse video microscopy was carried out as in E and distance migrated from the point of origin was quantified. (G) MCF10A cells overexpressing GLIS1 and p110 CUX1 alone or both were subjected to inverted Boyden chamber invasion assays. The percentage of cells able to migrate more than 20 µm into the matrix is shown for each line. The average of 3 independent experiments is shown. (H) Confocal Z-Stacks acquired 10 µm apart are aligned to demonstrate invasion of MCF10A-derivative cells into the matrix. Representative Z-stack for each condition is displayed. The leftmost image represents cells at the surface of the matrix (Depth = 0).

    Journal: Biology Open

    Article Title: Autocrine Activation of the Wnt/β-Catenin Pathway by CUX1 and GLIS1 in Breast Cancers

    doi: 10.1242/bio.20148193

    Figure Lengend Snippet: We established populations of MCF10A cells stably carrying lentiviral vectors that express nothing (vector), GLIS1, p110 CUX1, or both GLIS1 and p110 CUX1. (A) Wnt gene mRNA levels were measured by RT-qPCR. Und: Undetected. * P Value<0.05 on a Welch corrected T test when comparing the p110+Glis1 samples vs. each of the other 3. (B) β-Catenin protein levels were measured by immunofluorescence using antibodies specific for the active, non-phosphorylated form of the protein. Histogram shows the mean nuclear signal for β-Catenin for at least 50 cells per condition. * P Value<0.05 on a Welch corrected T test. A representative image of each condition is shown below. Scale bar: 20µm. (C) β-Catenin protein levels were measured by immunoblotting using antibodies specific for the active, non-phosphorylated form of the protein. γ-Tubulin is shown as a loading control. (D) MCF10A cells were transiently transfected with either the TOP of the FOP luciferase reporter, a vector expressing p110 CUX1 or an empty vector, and a vector expressing GLIS1 or the corresponding empty vector. Luciferase activity was measured in 3 independent transfections. * P Value<0.05 on a Welch corrected T test. (E) MCF10A cells ectopically expressing GLIS1, p110 CUX1 or both were imaged by time-lapse video microscopy and their speed of migration was quantified. The average of 3 independent experiments is shown with error bars representing standard error. ** P Value<0.01, *** <0.001 on 3 combined Welch corrected T tests using the Fisher method. (F) Time-lapse video microscopy was carried out as in E and distance migrated from the point of origin was quantified. (G) MCF10A cells overexpressing GLIS1 and p110 CUX1 alone or both were subjected to inverted Boyden chamber invasion assays. The percentage of cells able to migrate more than 20 µm into the matrix is shown for each line. The average of 3 independent experiments is shown. (H) Confocal Z-Stacks acquired 10 µm apart are aligned to demonstrate invasion of MCF10A-derivative cells into the matrix. Representative Z-stack for each condition is displayed. The leftmost image represents cells at the surface of the matrix (Depth = 0).

    Article Snippet: For TOP/FOP reporter assay, Cells plated at 8×10 4 per well in 12-well plates were cotransfected with 0.25 μg of either the TOP 8× or FOP 8× reporter plasmids (Addgene) and 0.5 μg of either CUX1 p110 or empty vector as effector.

    Techniques: Stable Transfection, Plasmid Preparation, Quantitative RT-PCR, Immunofluorescence, Western Blot, Control, Transfection, Luciferase, Expressing, Activity Assay, Microscopy, Migration

    We established populations of Hs578t cells stably carrying lentiviral vectors that express nothing (vector), GLIS1, p110 CUX1, or both GLIS1 and p110 CUX1. (A) Wnt gene mRNA levels were measured by RT-qPCR. Und: Undetected. * P Value<0.05 on a Welch corrected T test. (B) β-Catenin protein levels were measured by indirect immunofluorescence using antibodies specific for the active, non-phosphorylated form of the protein. Histogram shows the mean nuclear signal for β-Catenin for at least 50 cells per condition. * P Value<0.05 on a Welch corrected T test. A representative image of each condition is shown below. Scale bar: 20µm. (C) Hs578t cells were transiently transfected with either the TOP of the FOP luciferase reporter, and suboptimal amounts (50 ng) of effector plasmids coding for nothing (−), p110 CUX1 or GLIS1. Luciferase activity was measured in 3 independent transfections. * P Value<0.05 on a Welch corrected T test. (D) Hs578t cells ectopically expressing GLIS1, p110 CUX1 or both were imaged by time-lapse video microscopy and their speed of migration was quantified. The average of 3 independent experiments is shown with error bars representing standard error. ** P Value<0.01, *** <0.001 on a Welch corrected T test. (E) Time-lapse video microscopy was carried out as in D and distance migrated from the point of origin was quantified.

    Journal: Biology Open

    Article Title: Autocrine Activation of the Wnt/β-Catenin Pathway by CUX1 and GLIS1 in Breast Cancers

    doi: 10.1242/bio.20148193

    Figure Lengend Snippet: We established populations of Hs578t cells stably carrying lentiviral vectors that express nothing (vector), GLIS1, p110 CUX1, or both GLIS1 and p110 CUX1. (A) Wnt gene mRNA levels were measured by RT-qPCR. Und: Undetected. * P Value<0.05 on a Welch corrected T test. (B) β-Catenin protein levels were measured by indirect immunofluorescence using antibodies specific for the active, non-phosphorylated form of the protein. Histogram shows the mean nuclear signal for β-Catenin for at least 50 cells per condition. * P Value<0.05 on a Welch corrected T test. A representative image of each condition is shown below. Scale bar: 20µm. (C) Hs578t cells were transiently transfected with either the TOP of the FOP luciferase reporter, and suboptimal amounts (50 ng) of effector plasmids coding for nothing (−), p110 CUX1 or GLIS1. Luciferase activity was measured in 3 independent transfections. * P Value<0.05 on a Welch corrected T test. (D) Hs578t cells ectopically expressing GLIS1, p110 CUX1 or both were imaged by time-lapse video microscopy and their speed of migration was quantified. The average of 3 independent experiments is shown with error bars representing standard error. ** P Value<0.01, *** <0.001 on a Welch corrected T test. (E) Time-lapse video microscopy was carried out as in D and distance migrated from the point of origin was quantified.

    Article Snippet: For TOP/FOP reporter assay, Cells plated at 8×10 4 per well in 12-well plates were cotransfected with 0.25 μg of either the TOP 8× or FOP 8× reporter plasmids (Addgene) and 0.5 μg of either CUX1 p110 or empty vector as effector.

    Techniques: Stable Transfection, Plasmid Preparation, Quantitative RT-PCR, Immunofluorescence, Transfection, Luciferase, Activity Assay, Expressing, Microscopy, Migration